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Team:ZJU-China/labnote4.htm
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<h3>July 11</h3> | <h3>July 11</h3> | ||
<p>1. Identify FA and FB with PCR.</p> | <p>1. Identify FA and FB with PCR.</p> | ||
| - | <p><img src="https://static.igem.org/mediawiki/igem.org/a/a2/Zju_notebook3.jpg" width=" | + | <p><img src="https://static.igem.org/mediawiki/igem.org/a/a2/Zju_notebook3.jpg" width="500px"><p> |
<p>2. Amplify FB-7-10-plate bacteria in liquid LB medium.</p> | <p>2. Amplify FB-7-10-plate bacteria in liquid LB medium.</p> | ||
<h3>July 12</h3> | <h3>July 12</h3> | ||
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<p>2. Amplify FB-7-10-plate1, FB-7-10-plate2, FA-6.29-liq (DH5α), FA-lix and FB-lix bacteria for minipreparing plasmids later.</p> | <p>2. Amplify FB-7-10-plate1, FB-7-10-plate2, FA-6.29-liq (DH5α), FA-lix and FB-lix bacteria for minipreparing plasmids later.</p> | ||
<p>3. Identify FB with PCR again.We found there were problems with primers used in those reactions, thus leading to the failure.</p> | <p>3. Identify FB with PCR again.We found there were problems with primers used in those reactions, thus leading to the failure.</p> | ||
| - | <p><img src="https://static.igem.org/mediawiki/igem.org/9/9f/Zju_notebook4.jpg" width=" | + | <p><img src="https://static.igem.org/mediawiki/igem.org/9/9f/Zju_notebook4.jpg" width="500px"><p> |
<p>4. Ligation: D0 and pSB1C3, purified at July 08.</p> | <p>4. Ligation: D0 and pSB1C3, purified at July 08.</p> | ||
<h3>July 13</h3> | <h3>July 13</h3> | ||
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<p>Name: FB-7-14-5 Concentration: 33.1 ng/ul A260/280: 0.68</p> | <p>Name: FB-7-14-5 Concentration: 33.1 ng/ul A260/280: 0.68</p> | ||
<p>Name: FB-7-14-6 Concentration: 26.1 ng/ul A260/280: 1.64</p> | <p>Name: FB-7-14-6 Concentration: 26.1 ng/ul A260/280: 1.64</p> | ||
| - | <p><img src="https://static.igem.org/mediawiki/igem.org/3/3e/Zju_notebook6.jpg" width=" | + | <p><img src="https://static.igem.org/mediawiki/igem.org/3/3e/Zju_notebook6.jpg" width="500px"><p> |
<p>3. Amplify glycerol stock bacteria co-transformation (BL21*(DE3)) on 3 resistance plates, glycerol stock bacteria D0 on Amp plate.</p> | <p>3. Amplify glycerol stock bacteria co-transformation (BL21*(DE3)) on 3 resistance plates, glycerol stock bacteria D0 on Amp plate.</p> | ||
<p>4. Amplify FB-7-12-2, FB-7-12-4, FA-7-12-2-2 and FA-7-12-2-3 in LB liquid medium, named FB-7-14-2, FB-7-14-4, FA-7-14-2-2 and FA-7-14-2-3.</p> | <p>4. Amplify FB-7-12-2, FB-7-12-4, FA-7-12-2-2 and FA-7-12-2-3 in LB liquid medium, named FB-7-14-2, FB-7-14-4, FA-7-14-2-2 and FA-7-14-2-3.</p> | ||
<p>5. Digest FA-7-13-2-2, FA-7-13-2-3, FB-7-14-2 and FB-7-14-4 with Pst1.</p> | <p>5. Digest FA-7-13-2-2, FA-7-13-2-3, FB-7-14-2 and FB-7-14-4 with Pst1.</p> | ||
| - | <p><img src="https://static.igem.org/mediawiki/igem.org/d/de/Zju_notebook7.jpg" width=" | + | <p><img src="https://static.igem.org/mediawiki/igem.org/d/de/Zju_notebook7.jpg" width="500px"><p> |
<p>Maybe the concentration is too low that we can't recognize it.</p> | <p>Maybe the concentration is too low that we can't recognize it.</p> | ||
Revision as of 13:10, 25 September 2012
Week 4 (07.09-07.15): Make our own biobrick parts
July 09
Prepare liquid SOB medium for co-transformation.
July 10
Amplify glycerol stock bacteria FB (DH5α) on plates.
July 11
1. Identify FA and FB with PCR.
2. Amplify FB-7-10-plate bacteria in liquid LB medium.
July 12
1. Miniprep plasmid of pCJDFB from bacteria of July 06.
Name: FB-7-11 Concentration: 15.4 ng/ul A260/280: 1.88
2. Amplify FB-7-10-plate1, FB-7-10-plate2, FA-6.29-liq (DH5α), FA-lix and FB-lix bacteria for minipreparing plasmids later.
3. Identify FB with PCR again.We found there were problems with primers used in those reactions, thus leading to the failure.
4. Ligation: D0 and pSB1C3, purified at July 08.
July 13
1. Transformation:
No.name time of heat shock/s
1 D0-FA-FB 90
2 FA-FB 90
3 D0-FA-FB 30
4 FA-FB 30
5 ligation product 90
6 D0 90
2. Miniprep plasmid of FA and FB from bacteria of July 12.
Name: FA-7-13-1-1 Concentration: 15.8 ng/ul A260/280: 1.89
Name: FA-7-13-1-2 Concentration: 10.8 ng/ul A260/280: 2.06
Name: FA-7-13-1-3 Concentration: 13.6 ng/ul A260/280: 2.30
Name: FA-7-13-1-4 Concentration: 12.3 ng/ul A260/280: 2.13
Name: FA-7-13-2-1 Concentration: 9.4 ng/ul A260/280: 2.15
Name: FA-7-13-2-2 Concentration: 12.4 ng/ul A260/280: 1.94
Name: FA-7-13-2-3 Concentration: 20.1 ng/ul A260/280: 1.94
Name: FA-7-13-3-1 Concentration: 11.1 ng/ul A260/280: 2.23
Name: FA-7-13-lix Concentration: 13.3 ng/ul A260/280: 2.08
Name: FB-7-13-yy Concentration: 26.4 ng/ul A260/280: 1.84
July 14
1. Co-transformation:
No.name time of heat shock/s
1 D0-FA-FB 90
2 D0-FA-FB 30
Use SOC liquid medium.
2. Miniprep plasmid of FA and FB from bacteria of July 12.
Name: FB-7-14-1 Concentration: 42.9 ng/ul A260/280: 0.67
Name: FB-7-14-2 Concentration: 21.0 ng/ul A260/280: 2.69
Name: FB-7-14-3 Concentration: 44.7 ng/ul A260/280: 0.59
Name: FB-7-14-4 Concentration: 14.4 ng/ul A260/280: 2.09
Name: FB-7-14-5 Concentration: 33.1 ng/ul A260/280: 0.68
Name: FB-7-14-6 Concentration: 26.1 ng/ul A260/280: 1.64
3. Amplify glycerol stock bacteria co-transformation (BL21*(DE3)) on 3 resistance plates, glycerol stock bacteria D0 on Amp plate.
4. Amplify FB-7-12-2, FB-7-12-4, FA-7-12-2-2 and FA-7-12-2-3 in LB liquid medium, named FB-7-14-2, FB-7-14-4, FA-7-14-2-2 and FA-7-14-2-3.
5. Digest FA-7-13-2-2, FA-7-13-2-3, FB-7-14-2 and FB-7-14-4 with Pst1.
Maybe the concentration is too low that we can't recognize it.
July 15
1. Transformation:
No. name time of heat shock/s Strain
1 D0-FA-FB 90 DH10β
2 FA-FB 90 DH10β
3 D0-FA-FB 30 BL21*(DE3)
4 FA-FB 30 BL21*(DE3)
5 D0-pSB1C3 90 BL21*(DE3)
6 D0 90 BL21*(DE3)
2. Amplify bacteria in LB liquid medium:
No. Name From quantity
1 Co-7-15 co-transformation (BL21*(DE3)) 6
2 D0-7-15 D0-7-14 1
3 Co3-7-15-1-3-1 Co3-7-14-1 1
4 Co3-7-15-1-3-2 Co3-7-14-2 1
